Tuberculosis (TB) remains a global public health crisis despite being a curable disease. Access to the complete genomic sequence of the Mycobacterium tuberculosis (MTB) laboratory strain H37Rv and clinical isolate CDC1551 has facilitated investigations into the pathogenicity of MTB. However, mycobacterial factors that contribute to the virulence of MTB or modulate the interaction of this pathogen with the human host are only beginning to be elucidated. Despite previous studies suggesting that MTB has an extremely low mutation rate, there are compelling reasons to suspect that strain-specific attributes contribute directly to virulence and disease outcome. Our studies of infections in THP-1 cells with epidemiologically distinct clinical MTB strains provide strong evidence that specific clinical MTB strains may be differentially pathogenic. Clinical strains associated with TB outbreaks grow significantly faster in human macrophages than do non-outbreak strains. The rapid growth demonstrated by strains associated with outbreaks was highly correlated with rapid production of IL-10 and suppression of TNF?. These results suggest that the enhanced capacity of MTB to grow rapidly in human macrophages is a marker of virulence, and virulence of certain strains may be attributed to down-regulation of the Th1-type immune response. The following aims will address the hypothesis that differences in the genome or gene expression among clinical strains contribute to enhanced intracellular growth and/or modulate the macrophage response. Aim 1. To investigate genomic variation between the rapid-growth and slow-growth phenotypes of MTB. Comparative genomic analysis will be performed to identify genomic deletions associated with the respective growth phenotype. Clinical strains showing rapid or slow growth rates will be screened for deletions using PCR to identify deletions that are consistently associated with the growth phenotype. To determine if the phenotype is a consequence of specific gene interruptions, we will complement selected deletions and test the complemented strains in the THP-1 infection model. Aim 2. To identify genes differentially expressed in MTB clinical strains exhibiting enhanced intracellular growth. The role of disrupted or deleted genes in the intracellular growth rate and/or modulating the cytokine response will be further investigated by measuring transcription levels during the course of THP-1 infection using RT-PCR. If deletions are not readily identified as being associated with the growth rate, microarray analysis will be used to examine global gene expression. The ability to combine genomic information with pathogenesis studies employing diverse clinical strains will enable us to continue to unravel the molecular basis of MTB virulence. Furthermore, having the ability to identify relevant strain characteristics could potentially impact how we treat and control the spread of TB. PUBLIC HEALTH RELEVANCE: Little is understood about strain-specific properties of clinical isolates of Mycobacterium tuberculosis, the causative agent of tuberculosis (TB). We plan to study clinical isolates in human macrophages to identify microbial factors that are involved in virulence. Identification of relevant strain characteristics could potentially impact how we treat and control the spread of TB. Furthermore, we anticipate that our results will elucidate whether or not strain virulence will impact the protection afforded by a new candidate vaccine.